Nanomaterials possess unique physical properties due to their confinement down to the nanometer scale in at least one dimension. Combination of nanomaterials and biological processes opens up new horizons for modern biotechnological applications, particularly for biosensing. Not only nanomaterials enable miniaturisation of devices, but also ensure their high sensitivity due to the significantly increased surface-to-volume ratio.
As nanomaterials in general, nanowires and their arrays in particular, confined down to the nanometer thickness, show many advantages over conventional wires, such as significantly increased signal-to-noise ratio, increased charge carrier transport and low detection limit. Therefore, the nanowires are now considered a new class of ultrasensitive electrochemical tools and electrochemical sensors.
However, since these sensor devices operate in ultralow current regimes and are in general very sensitive to various influences from side parameters, which generally decreases the stability and reliability of the sensor operation, special designs and array layouts are advised, which enable maximum control of the sensors performance. This includes a gate voltage control from front side via a stable electrochemical reference electrode system and from back side via a back gate contact. In addition a shift to the frequency domain for alternative sensor readout stabilisation aids more reliable performance in biosensor applications.
Silicon nanomaterials are a type of nanomaterials with attractive properties, including excellent semiconducting, mechanical, optical properties, favourable biocompatibility, surface tailorability, and are relatively compatible with conventional silicon technology. A silicon nanowire (hereinbelow, “SiNW”) is defined as a “one-dimensional” silicon nanostructure exhibiting a length-to-diameter ratio of 1000 or more. The diameter of a regular SiNW is in order of manometers. At this scale, quantum mechanical laws become totally dominant, and SiNWs have many interesting properties that are not seen in larger objects and three-dimensional materials. Since quantum confinement of electrons in SiNWs to one dimension is predicted to be substantial only at diameters below 3 nm, the band structure is strongly modified for these nanowires having diameters of manometers. The band gap in SiNWs increases for smaller diameters and a direct band gap can be obtained for sufficiently small diameters. Silicon nanowires have been extensively explored for myriad applications ranging from electronics to biology (M. Zhang et al 2008).
A field-effect transistor (FET) is a semiconductor-based device with relatively low power consumption used for switching or signal amplification in electrical circuits. Widely used transistors are metal oxide semiconductor field-effect transistors (MOSFETs) having two highly doped domains (n- or p-type) separated by a p- or n-type domains, respectively, with p-n or n-p junctions at the interface enabling a tunnelling operation. Common elements for doping of silicon are boron (p-type) or arsenic/phosphoric (n-type).
The doped domains are connected to source (S) and drain (D) ohmic contacts. A metal gate is placed on top of the area between the S and D contacts, separated by a thin insulating oxide layer. Conductance of the transistor is controlled by an external field, for instance the gate field. By applying a positive or negative gate voltage at the gate, electrons are either attracted or repulsed at the semiconductor-oxide interface. Once the threshold voltage is reached, an inversion layer forms a conducting channel between the n- or p-type regions connected to the S and D contacts. Size of the formed channel depends on both, the external gate field and the selection of the materials. Applying an additional SD bias voltage enhances the charge carrier movement through the channel and narrows the channel at one certain end. Thus, conductance is altered, and electrons become attracted by the drain.
Ion-sensitive FETs (ISFETs) are similar to the common MOSFET, but the metal gate electrode is replaced with an electrolyte solution carrying analyte molecules and containing a reference electrode. The electrolyte solution is defined as a “liquid gate” which controls the current between the S and D contacts. The liquid gate electrode is separated from the channel by the gate insulator and/or an ionic “double layer” barrier, which is sensitive, for example to protons, and therefore, suitable for pH measurements. In general, binding reactions of charged analytes with corresponding ligand groups at the ISFET sensor surface cause a surface charge which leads to an additional surface potential. This change in surface potential is monitored and can be related to the number of adsorbed analyte molecules. Using different surfaces and surface functionalisation techniques, sensitivity to specific target analyte molecules can be achieved with this sensor. For sensing, it is important to include a reference electrode to control the solution potential and to apply the liquid gate voltage.
Conductance in ISFETs is strongly dependent on the charge density at the oxide-electrolyte interface. ISFETs are sensitive towards any electrical fields in general, and to charged molecules in particular. Depending on the point of zero charge of the surface and the pH of the electrolyte a respective surface charge is building up, the charging of the terminal surface groups behaves like a local gate potential. Therefore, it is possible to detect variations in the pH of the solution, i.e. taking or giving away protons at the functional interface is sensed. For the transfer characteristics of the devices, the source drain current (ISD) is measured as a function of the applied gate voltage VG. In a semi-logarithmic scale, the steepest slope of the curve ISD vs VG is defined as a sub-threshold swing with the threshold voltage VT at which the current is switched on due to the formation of an inversion layer at the insulator-semiconductor interface. At this point the FET switching can happen at maximum speed. By analysing this slope, which is constant over several orders of magnitude, curve shifts at a fixed current due to attached charged molecules can be determined (ΔVT).
Because of the possible (de)protonation of the surface oxide, pH measurements are usually applied to probe the device performances for potential application in biosensing. By changing pH, the surface potential is directly affected and influences the VT. As a result, the curve of ISD vs VG shifts to lower or higher voltage values. Sensitivity of ISFETs is thus determined by the maximum possible shift because of a pH change, and defined by the Nernst limit at 300 K, as explained in detail by Tarasov in his PhD thesis “Silicon Nanowire Field-effect Transistors for Sensing Applications” (2012).
As most biomolecules display charges at their outer surface, ISFETs are equally sensitive to biomolecules such as proteins or DNA. Thus, their ISD vs VG curve shifts parallel to the VG X-axis when the charge density changes, and can be further analysed the same way as described for the aforementioned pH measurements.
Bavli et al (2012) showed a two-dimensional (planar) FET (used as a molecularly controlled semiconductor resistor), run in liquid environment, for the detection of different analytes on a lipid bilayer functionalized surface with a detection limit in the μg/ml range, which clearly indicates that this FET lacks low detection limits. SiNWs are integrated in field-effect transistors (FETs) to build sensor devices with strong signal amplification at low power consumption, which is advantageous for portable or implantable devices. One-dimensional FETs strongly benefit from an extreme surface area to volume ratio, which allows effective channel gating from even just a few adsorbed analyte molecules. SiNW-based FET sensing was described first by Cui et al in 2001 using vapour-liquid-solid (VLS) grown silicon nanowires (SiNWs) for pH sensing and detecting the binding of streptavidin protein on biotin-labelled wires. To date they demonstrated sensing of DNA/PNA hybridisation (Hahm and Lieber 2004), viruses down to single virus detection events (Patolsky et al 2004), using antibodies as receptors, multiplexed sensing (Patolsky et al 2006) and cell potentials (Jiang et al 2012).
SiNW-based FETs used for biosensing belong to the group of ISFETs and therefore operate in liquid medium. SiNWs compared to ISFETs offer higher surface-to-volume ratios for better sensitivity and small cross-sectional conduction pathways. Thus, they can overcome the detection limits of planar ISFETs. SiNW-based FETs have a conductance of 4-10 times higher than planar standard ISFETs of the same sizes due to their high surface-to-volume ratio and the efficient penetration of the gate field.
In biosensing applications of SiNW FETs, as in ISFETs, liquid also acts as a gate electrode and variations in the surface potential are converted to a conductance change in the channel. Actually, different from planar ISFETs, the one-dimensional SiNWs themselves are conduction channels, which are fully affected by the surface potential. Biological recognition events strongly alter the surface potential of the SiNWs even at low analyte concentrations. Charged biomolecules can locally act as liquid gate leading to resistivity changes, which (resistivity) is very sensitive to the biorecognition event. Due to signal amplification by the FET, the signal can be detected by significant jumps in the voltage at a fixed source drain current. Commonly, single-crystalline nanowires are with p- or n-type doping containing charge carriers that are attracted or repelled by the attaching charged biomolecules.
SiNWs can be combined with existing processing technology on silicon wafers to fabricate chips, which significantly reduces the cost of disposable chips. Many groups developed SiNW arrays in top-down fabrication to measure biorecognition events like protein detection using antibodies (Elnathan et al 2012) or hybridisation (G. Zhang et al 2008). In order to use these devices as biosensors, the silicon dioxide (SiO2) surface of SiNWs should be functionalised with biorecognition elements in a controllable manner.
For all the different applications, interface engineering and chemical functionalisation of the transducer (SiO2) surface are crucial for biosensor development to assure an excellent sensor performance. Requirements are a stable receptor attachment under varying conditions, while preserving the functionality of receptors at the same time. Additionally, binding strategies will enhance the receptor orientation towards the target in solution whereas blocking protocols try to avoid unspecific attachment to reduce background signals. The main focus here is on chemical functionalisation of silicon dioxide surfaces of SiNWs and evaluation of the receptor-analyte interactions with the immobilised receptors using optical and electrical sensing methods using the SiNW-based FET.
Real-time, label-free, portable, low-cost, flexible and reliable sensors with lab on chip systems are long-needed biomedical diagnostic devices, and they are still challenging. Further requirements to evaluate novel biosensors are their sensitivity to detect low levels of the analyte, selectivity to avoid false positive signals and a fast response time that allows for rapid diagnosis. In terms of fabrication, the integration into existing technologies and production, the versatility of application and the possibility to produce at low costs are crucial. Reversibility of the biosensor would allow for repeated measurements to improve comparability.
Integration of nanomaterials into biosensor's transducer is one of the possibilities to meet some of the above listed requirements especially in terms of miniaturisation and sensitivity due to their high surface-to-volume ratio and the confinement within the nanometer scale in at least one dimension which leads to changes in the physical properties. SiNW based FETs fulfil many of these requirements. The use of nanomaterials and the possible integration into Si wafer processing technology enables miniaturisation of devices at low costs. The immediate current jump upon physicochemical changes without need for analyte labelling meet the requirements for the modern biosensor of being real-time and label-free.
J.-H. Ahn et al (2014) described a SiNW FET with local side-gates and SiNW surface decorated with palladium nanoparticles for hydrogen detection. The drain current of their sensor reversibly responded to hydrogen concentration. A control experiment, which used the sensor with a bare SiNW without Pd nanoparticles, proved that the hydrogen-sensitivity is originated from the palladium nanoparticles decorating the SiNW surface. S.-W. Ryu et al (2010) discussed the SiNWs decorated with gold nanoparticles for use in a biosensor for label-free DNA detection with enhanced sensitivity. The electric current flow between a source and a drain electrode was measured in order to sense the immobilisation of tested oligonucleotides and their hybridisation with target oligonucleotides.
S. Yan et al (2010) demonstrated the fabrication method of the SiNWs decorated with gold nanoparticles. These SiNWs were directly fabricated into an electrode for detection of bovine serum albumin in solution without any pre-treatment. H. Huang et al (2014) described a method for producing a finely dispersed CuO nanoparticle electro-catalyst on a SiNW surface by combining metal-assisted chemical etching, electroless deposition, and thermal oxidation. They used the sensor based on the SiNWs decorated with CuO for sensing hydrogen peroxide.
To improve stability of the sensor arrays and ensure reproducibility of the readout, sensors of disclosed embodiments are fabricated in a parallel batch process on standard silicon wafers. To enable a control in bioassays, reference sensor structures such as temperature sensors, pH sensors and ionic strength sensors are added to the sensor chip. As discussed below, the design of the pixel array in a three-electrode configuration including a reference electrode and a counter electrode enables an additional operation of the optoelectronic sensor pixel in the frequency domain and helps to stabilize the electronic readout when recording very small DS current changes. Therefore this sensor can be additionally used for impedance spectroscopy applications. A combination of potentiometric and impedimetric readout enables a more reliable sensing of biomolecules with the potential to sense beyond the Debye screening of electrical charges in an electrolyte solution, which is usually the limiting factor in SiNW sensors with only potentiometric or conductometric readout.
Various embodiments may allow various benefits, and may be used in conjunction with various applications. The details of one or more embodiments are set forth in the accompanying figures and the description below. Other features, objects and advantages of the described techniques will be apparent from the description and drawings and from the claims.